No abstract

The complete conjugal transfer gene region of the IncW plasmid R388 has been cloned in multicopy vector plasmids and mapped to a contiguous 14.9-kilobase segment by insertion mutagenesis. The fertility of the cloned region could still be inhibited by a coresident IncP plasmid. The transfer region has been dissected into two regions, one involved in pilus synthesis and assembly (PILw), and the other involved in conjugal DNA metabolism (MOBW). They have been separately cloned. PILw also contains the genes involved in entry exclusion. MOBW contains oriT and the gene products required for efficient mobilization by PILW. MOBw plasmids could also be mobilized efficiently by PILN, the specific pilus of the IncN plasmid pCUl, but not by PILp, the specific pilus of the IncP plasmid RP1.

No abstract

In the presence of the Tn21 transposase, plasmids that contain a single Tn21 inverted repeat sequence fuse efficiently with other plasmids. This reaction occurs in recA strains, is independent of the transposon-encoded resolution system, and results in insertions into different sites in the recipient plasmid. All fusion products studied contained at least one complete copy of the donor plasmid; most also contained some duplication of it as well. The data are consistent with processive models of transposition.

SummaryThe conjugative transfer region of the IncX plasmid R6K (TRA X ) was analysed by transposon mutagenesis and DNA sequencing. Tn5 tac1 insertional mutations localized TRA X to a 14.8 kb segment containing the ␣ origin of transfer (oriT␣), genes involved in conjugative DNA-processing (Dtr X ) and genes involved in pilus synthesis and assembly (Mpf X ). A second functional oriT, oriT ␤, was located at a distance of 5.3 kb from oriT␣ and was outside TRA X . Mpf X occupied a segment of 10 kb, as judged by the location of insertions conferring resistance to infection by the X pilusspecific phage X-2. At both sides of Mpf X there were insertions that were Tra ¹ but X-2 sensitive, suggesting that the mutations were in Dtr X . This region was sequenced and three genes were identified: taxA, taxB, and taxC. The overall organization was oriT␣-taxA-taxC-Mpf X -taxB. taxC coded for a oriT-relaxase that belongs to the VirD2 family. taxA coded for a protein of 181 amino acids that showed similarity to TraY of F-like plasmids and to the Arc-repressor superfamily. TaxB showed similarity to TraG-like proteins, a protein superfamily probably involved in coupling the relaxosome to the DNA-transport apparatus. TaxA and TaxC are required for oriT nicking in vivo. The nicking reaction was mistakenly assumed by Flashner et al. (1996) to represent a feature of the vegetative replication origins. However, insertions or deletions disrupting taxA and taxC affected conjugation but not replication of R6K. Conversely, protein , which is absolutely required for replication of R6K, was not required for conjugative transfer. In addition, protein DDP3, which is also assumed to have a role in replication, was found to be a positive modulator of bacterial conjugation. Taken together, these results rule out a direct and essential involvement of conjugation proteins in R6K vegetative replication, and also rule out the requirement of replication protein for conjugation.